FIG 5.

miR-155 interacts with the rs61764044 major allele and acts together with miR-425 to further suppress NPPA mRNA and ANP protein levels in human cardiomyocytes. (A and B) Allele-specific effects of miR-425 plus miR-155 (A) and anti-miR-425 plus anti-miR-155 (B) on the luciferase activity produced by the WT-Luc and rs61764044 Minor-Luc constructs in COS-7 cells. The rs61764044 Minor-Luc construct contains the minor alleles of rs61764044 and rs5068. (C) NPPA mRNA levels in hESC-CMs transfected with the indicated miRNAs relative to scrambled negative-control miRNA mimic. (D) Nt-proANP protein levels in the medium of hESC-CMs transfected with the indicated miRNAs relative to scrambled negative-control miRNA mimic. For miR-425 plus miR-155 cotransfection, the total amount of transfected miRNA was held constant under all experimental conditions (10 nM total concentration in COS-7 cells, 100 nM total concentration in hESC-CMs). For the conditions labeled with “miR-425” or “miR-155,” the total concentration of transfected miRNA was held constant by the addition of the scrambled negative-control miRNA mimic. For anti-miR-425 plus anti-miR-155 cotransfection, the total amount of transfected anti-miRNA was held constant under all experimental conditions (200 nM total concentration in COS-7 cells). For the experimental conditions labeled with “anti-miR-425” or “anti-miR-155,” the total concentration of transfected miRNA was held constant by the addition of the scrambled negative-control anti-miRNA. Data are means ± SEMs (n = 6 biological replicates). P values are from a two-tailed independent t test. *, P < 0.001 versus miR-425, and #, P < 0.001 versus miR-155, by one-way ANOVA with Bonferroni post hoc testing.